U.S. patent application Publication No. US 2003/0070939 of Walters et al, published Apr. 17, 2003, which was ultimately based upon PCT International Application PCT/US97/09300, of Walters et al, filed Jun. 1, 1997, and published as PCT International Publication Number WO 98/56893 on Dec. 17, 1998, discloses a method and apparatus for treating materials with electrical fields having varying orientations. In FIG. 13, of both the originally filed PCT application and the published PCT application, an array of eight in vitro test wells is disclosed. As disclosed on page 54 of the PCT International Publication Number WO 98/56893, each test well can be pulsed in a desired way either different from or the same as other test wells. Moreover, on page 53 of the PCT published application, there is a disclosure that generally, N applied pulses can be routed to N groups of electrodes in sequence. Furthermore, successive groups of electrodes in the N groups of electrodes can be comprised of different individual electrodes. Alternatively, successive groups of electrodes in the N groups of electrodes can be comprised of the same individual electrodes. Clearly, the disclosures in PCT International Publication Number WO 98/56893, published on Dec. 17, 1998, provide high throughput of electroporation tests or treatments. The array of test wells in FIG. 13 can be regarded as either a matrix array having one row and eight columns, or, alternatively, a matrix array having eight rows and one column.
It is noted that the disclosures in PCT International Publication Number WO 98/56893 are contained in U.S. Pat. No. 6,117,660 of Walters et al, which is incorporated herein by reference. U.S. Pat. No. 6,117,660 was issued on Sep. 12, 2000.
U.S. Pat. No. 6,352,853 of King et al, issued Mar. 5, 2002, based upon a utility patent application filed Nov. 16, 1999, which was related to a provisional patent application filed Dec. 7, 1998, discloses multi-channel electrode arrays used in electroporation. U.S. Pat. No. 6,352,853 discloses that multi-channel electrode systems are used for high throughput introduction of exogenous molecules into cells, or to avoid the need for transferring cells from culture containers to electroporation cuvettes. A multi-channel electroporation apparatus includes a plurality of pairs of electrodes positioned in respective ones of a plurality of chambers that hold the exogenous materials and the cells. U.S. Pat. No. 6,352,853 discloses that multi-channel electroporation devices contain 8 or 96 pairs of coaxial electrodes (Genetronics, Inc., San Diego, Calif.). These devices are used for electroporation in standard 96-well plates, which consist of 8 rows and 12 columns of wells and have a standard size of about 8.5 (W) cm.times.12.7 cm (L), with a standard center-to-center spacing of 9.0 mm between wells. For a standard 96 well plate, there are 96 wells, 96 pairs of electrodes (one pair of electrodes for each well), 96 first electrode conductors, and 96 second electrode conductors. Thus, there are 192 first or second electrode conductors to connect to 96 pairs of electrodes for 96 wells. The complexity involved with 192 first or second electrode conductors for a standard 96 well plate is very high. Moreover, the electronic apparatus that is needed to connect to and drive the 192 first or second electrode conductors is also very high. In this respect, it would be desirable to provide an electroporation apparatus that considerably reduces the complexity with respect to 192 first or second electrode conductors and the complexity with respect to the electronic apparatus needed to connect with and drive 192 first or second electrode conductors for a standard 96 well plate used for electroporation. More generally, it would be desirable if an electroporation apparatus were provided in which the number of conductors connected with electrodes for electroporation were considerably less than the number of electrodes.
Parenthetically, it is believed by the inventors of this present patent application that the Genetronics devices referred to in U.S. Pat. No. 6,352,853 have been superseded by BTX Products discussed herein below.
Further with respect to U.S. Pat. No. 6,352,853, electrodes are disclosed wherein each electrode includes a 24 electrode teeth. Each electrode is connected either to a positive (+) electric potential or to a negative (−) electric potential. Each row of 24 wells in a standard 384 well plate receives one 24 teeth electrode connected to a positive (+) electric potential and one 24 teeth electrode connected to a negative (−) electric potential. With the electroporation system provided in U.S. Pat. No. 6,352,853, a full row of 24 wells is treated simultaneously. There are 16 rows of 24 wells. Therefore, there are 32 electrodes needed to accommodate the 16 rows. Therefore, U.S. Pat. No. 6,352,853 has found a way to reduce the number of electrodes for treating all of the wells in a 384 well plate from 768 conductors to 32 conductors.
Yet, with U.S. Pat. No. 6,352,853, there is no way to electrically treat any one of the 24 wells in a full row of 24 well any differently from any other well in the full row of 24 wells. Since there are 16 rows of wells in a standard 384 well plate, only 16 different modalities of electrical treatment can be applied to the 384 wells. Stated differently, with U.S. Pat. No. 6,352,853, the individual wells are not individually addressable. To provide greater versatility, it would be desirable to provide an electroporation apparatus in which 384 electrode pairs in all 384 wells in a standard 384 well plate can be treated with a different electrical modality without the need for 768 conductors for connection with 364 pairs of electrodes. More generally, it would be desirable if an electroporation apparatus were provided in which pairs of electrodes in all wells in a standard well plate can be treated with a different electrical modality without the need for a pair of electrical conductors for each electrode pair. Stated somewhat differently, it would be desirable if individual wells in a standard well plate could be individually addressable.
With respect to BTX Products, a printout of ten pages from an Internet web site for BTX Products, having a URL of http://www.btxonline.com, which were printed on Dec. 16, 2002, include a three page description of HT-3000 High Throughput Electroporation Generator, a one page image of HT-3000, a two page description of Electroporation Plate Handlers, a one page image of HT-200, a one page description of Electroporation Plates, and a one page description of Transfection Optimization Software. The BTX Electroporation Plates use gold-plated, disposable multi-well electroporation plates which are used with high throughput electroporation apparatus. Such BTX gold-plated, disposable multi-well electroporation plates are quite expensive, considering the effort and expensive materials used to produce them. In essence, these BTX gold-plated, disposable multi-well electroporation plates provide both multiple electrodes and multiple wells in an integrated unit. Therefore, when the BTX gold-plated, disposable multi-well electroporation plates are disposed of, both the multiple electrodes and the multiple wells are disposed of at the same time. In this respect, it would be desirable for an electroporation apparatus to be provided in which multiple electrodes are not disposed of when multiple wells are disposed of.
On the other hand, simple, plastic disposable multi-well plates are well known and are much less expensive to make and use than the BTX gold-plated, disposable multi-well electroporation plates. Yet, it is not known that such well known simple, plastic disposable multi-well plates are used as wells in which multiple electroporations are carried out. In this respect, it would be desirable for an electroporation apparatus to be provided in which simple, plastic disposable multi-well plates can be employed and disposed of readily.
Returning to U.S. Pat. No. 6,352,853, this patent discloses a number of ways in which the electrode combs are supported. Yet, in each way of electrode comb support, once the electrode combs have been placed in the respective wells of a standard well plate, access to the respective wells is prevented by the support structure for the electrode combs. There may be times when it would be desirable to access the respective wells of a standard well plate even after electrodes have been placed in the respective wells. Generally, therefore, it would be desirable to provide a support for electrodes that are placed in the wells of a standard well plate wherein the electrode support allows access to the wells with the electrodes positioned in the wells.
Each well in a standard 96 rectangular well plate has a predetermined well volume. When pairs of rectangular electrodes are placed into the wells, the predetermined volumes are reduced, thereby leaving a reduced volume for conducting electroporation of materials. To assure that the predetermined volumes are reduced to a minimum by the electrodes so that the remaining volumes available for electroporation are at a maximum, it would be desirable if pairs of rectangular electrodes were placed into the rectangular wells so that the rectangular electrodes closely fit against adjacent walls of the wells.
Thus, while the foregoing body of prior art indicates it to be well known to use a matrix array of pairs of electrodes for a well plate which includes a matrix array of wells, the prior art described above does not teach or suggest a multiple electrode pair array which has a two-dimensional matrix array of electrode conductors and which has the following combination of desirable features: (1) provides an electroporation apparatus that considerably reduces the complexity with respect to 192 first or second electrode conductors and the complexity with respect to the electronic apparatus needed to drive 192 first or second electrode conductors for a 96 well plate used for electroporation; (2) provides an electroporation apparatus in which the number of conductors connected with electrodes for electroporation is considerably less than the number of electrodes; (3) provides an electroporation apparatus in which 384 electrode pairs in all 384 wells in a 384 well plate can be treated with a different electrical modality; (4) provides an electroporation apparatus in which all pairs of electrodes in all wells in a well plate can be treated with a different electrical modality without the need for a pair of electrical conductors for each electrode pair; (5) provides an electroporation apparatus in which multiple electrodes are not disposed of when a multiple well plate is disposed of; (6) provides an electroporation apparatus in which simple, plastic disposable multi-well plates can be employed and disposed of readily; (7) provides a support for electrodes that are placed in the wells of a standard well plate wherein the electrode support allows access to the wells when the electrodes are positioned in the wells; and (8) provides that pairs of rectangular electrodes are placed into the rectangular wells of a multi-well plate so that the rectangular electrodes closely fit against adjacent walls of the wells. The foregoing desired characteristics are provided by the unique multiple electrode pair array of the present invention as will be made apparent from the following description thereof. Other advantages of the present invention over the prior art also will be rendered evident.